The most abundant post-translational modification of proteins, glycosylation, remains practically unexplored to date at the proteome scale because of a dearth of methods for profiling the complex glycoproteome. Glycosylation of proteins plays an important role in many biological processes, including, for example, cell signaling, cell-cell interactions and the immune response. Further, the majority of protein-based biopharmaceuticals approved or in clinical trials bear some form of post-translational modification (PTM), which, in some cases, can profoundly affect protein properties relevant to their therapeutic application. A better understanding of the biological functions of glycosylation will facilitate the engineering of next-generation protein and peptide therapeutics with glycosylation profiles optimized for the respective therapeutic approach.
Analyzing the glycoproteome is technically challenging, because, as a post-translational process, glycosylation is non-templated, and, unlike other post-translational modifications (e.g., phosphorylation or methylation), glycan structures found on glycosylated proteins are highly complex. A single protein can have tens or hundreds of different glycan attachments, and glycosylated forms of proteins are often found in low abundance in the cell. The very different chemistries of proteins and glycans present additional challenges in applying analytic methods, such as mass spectrometry, in the glycoproteomic context, and current methods for isolating and/or separating glycoproteins for analytical processing lack in performance. See Doerr, Glycoproteomics Nat Meth 2012, 9(1):36, and Walsh et al., Post-translational modifications in the context of therapeutic proteins Nat Biotech 2006 24(10).